We will test the idea that there is a class of messenger RNA species whose synthesis is subject to stringent control, and another class whose synthesis is not. To do so, we will measure the functional levels of a variety of messengers in amino acid starved, rel plus and rel minus cells. We have previously demonstrated the occurrence of aberrant translation in rel minus cells. We propose to identify some of the specific amino acid substitutions which occur, and to study misreading of synthetic polynucleotides in vitro by rel minus ribosomes. We will also test the hypothesis that ribosome binding of uncharged tRNA competes with the binding of illegitimate charged species, and that rel minus ribosomes suffer a reduced affinity for uncharged tRNA. We will also test the idea, suggested by previous observations, that rel minus ribosomes sometimes terminate prematurely during amino acid starvation. We have identified a new gene (spoT) whose product is involved in the metabolism of the MS nucleotides. We propose to: (1) examine the effect of spoT genotype on various controversial aspects of the stringent response; and (b) identify and isolate the spoT gene product, in order to study the factors which regulate its activity. We have shown that protein synthesis is subject to stringent control by ppGpp in whole cells. We propose to find out whether this inhibition pertains to peptide chain initiation, elongation, or both. We have observed marked changes in the levels of three novel nucleotides when a eukaryotic organism (yeast) is subjected to conditions where RNA accumulation is restricted. We propose to characterize these nucleotides, and to find out what they have to do with control of RNA accumulation.